The disclosed compounds generally relate to the field of dithio reagents. In particular, the disclosed compounds generally relate to fluorescent dithio reagents that are useful for detecting thiol-containing compounds.
Thiols are ubiquitous in cellular biochemistry, playing important roles in determining protein structure (as disulfide linkages) and enzymatic mechanisms (as covalent catalysts). Furthermore, the redox state in the cell is largely regulated by the thiol/disulfide status of glutathione in the cell (i.e., GSH vs GSSG). In addition, reduced glutathione (i.e., the thiol form or GSH) also plays a control role in drug metabolism by attacking electrophilic atoms. Therefore, thiol detection and quantitation is important in cellular biochemistry, and to date has been accomplished most commonly by performing UV-Visible assays using calorimetric reagents such as Ellman's reagent.
Fluorescence detection systems (e.g., fluorescence spectroscopy) have been widely used to study the structure, mechanism and function of different proteins and enzymes, and especially in enzymatic activity or binding assays. Fluorescence detection systems are useful in that they generally have high sensitivity and a good dynamic range for detection. In addition, many generic fluorescent reagents are available, as well as commercially available equipment for detecting particular reagents. Fluorescence detection systems may be amenable to high throughput screening (e.g., using any bench-top fluorescence plate reader). Potential drawbacks associated with some fluorescence reagents may include photobleaching, stability, and purity of the utilized fluorophore. Some commercially available fluorescent labeling reagents are mixtures of isomers or have high photobleaching or cause uncontrolled labeling, which prevents them from giving reliable and reproducible results.
As such, fluorescent dithio reagents are desirable. In particular, fluorescent dithio reagents that are photostable, single isomers are desirable.